Treatment of LFA-1 associated disorders with increasing doses of LFA-1 antagonist

ABSTRACT

A method is provided for reducing the occurrence of fever, headache, nausea and/or vomiting associated with administration of a therapeutic compound to a mammal in need thereof, comprising administering to the mammal a first conditioning dose of a non-target cell depleting compound which binds to a cell surface receptor on a target mammalian cell; and administering a second therapeutic dose of the compound, wherein the second dose is higher than the first dose.

This is a continuation under 37 CFR §1.53(b) of co-pending U.S.application Ser. No. 10/655,905, filed Sep. 5, 2003, which is adivisional of U.S. application Ser. No. 09/936,603, filed Feb. 11, 2002,which claims priority to international application no. PCT/US00/07189,filed Mar. 17, 2000, which claims the benefit under 35 USC §119(e)(1) toprovisional patent applications Ser. No. 60/125,351 and Ser. No.60/125,228, both filed Mar. 19, 1999, the disclosures of which areincorporated by reference herein in their entirety.

FIELD OF THE INVENTION

The invention relates to methods of treating mammals, for examplehumans, to reduce the occurrence of undesired administration reactions,to treat an LFA-1 mediated disease, to condition a mammal to toleratehigh doses of a therapeutic compound and to down modulate a cell surfacereceptor.

DISCUSSION OF THE BACKGROUND

Administration of many therapeutic agents rapidly induces adverse sideeffects, or events, including but not limited to fever, headache,nausea, vomiting, breathing difficulties and changes in blood pressure.These adverse events limit the amount of a drug or therapeutic compoundthat can be given, which in turn limits the therapeutic effectivenessthat could be achieved with higher doses of the drug. There is acontinuing need to develop techniques which limit the toxicity of higherdrug doses so that therapeutic efficacy can be improved. This needexists for both polypeptide and non-polypeptide compounds.

Antibodies are one type of polypeptide compound for which there arefrequently adverse events upon administration which limit the dose ofthe compound that can be administered. One compound associated withadverse side effects is the murine monoclonal antibody OKT3. OKT3 bindsto the CD3 protein complex that is associated with the T cell receptor(TCR) found on the surface of all T lymphocytes. Administration of OKT3to humans rapidly reduces the number of circulating T cells (e.g. OKT3is a cell depleting compound) and reduces the amount of cell surface TCRfound on those T cells that remain (Cosimi, et al., 1981 N EngI J Med,305(6), 308-314). The immunosuppressive effects of OKT3 have beentherapeutically useful in the treatment of renal transplant rejection(Goldstein & Group, 1985 M Engl J Med, 313(6), 337-342). However,administration of OKT3 induces a number of adverse side effects,including fever, chills, nausea, vomiting and tightness of chest. Theseside effects are believed to be caused by cytokine release from T cellsdue to OKT3-induced activation (Abramowicz, et al., 1989Transplantation, 47(4), 606-608) and complement activation (Raasveld, etal., 1993 Kidney International, 43 1140-1149).

Several strategies have been developed to reduce the OKT3-induced sideeffects. Anti-inflammatory steroids have been shown to attenuate theOKT3-induced cytokine release (Goldman, et al., 1989 Lancet, ii (8666),802) (Chatenoud, et al., 1990 Transplantation, 49(4), 697-702), andindomethacin can reduce the febrile response (First, Schroeder,Hariharan, Alexander, & Weiskittel, 1992 Transplantation, 53 (1),91-94). A standard 5 mg dose of OKT3 administered as a 2 hour infusioninstead of the usual bolus injection was better tolerated and reducedcomplement activation, but not the cytokine release (ten Berge,Buysmann, van Diepen, Surachno, & Hack, 1996 Transplant Proc. 28 (6),3217-3220). The adverse events induced by OKT3 are most significantafter the first dose. While the initial dose (typically 5 mg) inducescytokine release and activates complement, it also eliminates the targetT cells. With fewer T cells and reduced TCR density on those that doremain, subsequent doses of OKT3 induce less cytokine release(Chatenoud, et al., 1989 N Engl J Med, 320 (21), 1420-1421). One groupfound that after four daily doses of 5 mg, dosing could safely beescalated to 10, 15, and 25 mg over the next 3 days (Woodle, et al.,1996 Clin Transplantation, 10, 389-395).

Adverse events have also been associated with the initial administrationof monoclonal antibodies directed to other cell surface molecules. Ahumanized anti-CD4 monoclonal antibody induced fever, chills,hypotension and chest tightness when given intravenously to psoriasisand rheumatoid arthritis patients (Isaacs, et al., 1997 Clin ExpImmunol, 110, 158-166). This treatment down-modulated expression of CD4and caused a reduction in the number of circulating CD4-positive Tcells, and but was not completely depleting. Bispecific antibodies thatinteract with the CD64 molecule, a receptor for the constant region ofimmunoglobulin (Fc gamma R1), and tumor associated molecules (epidermalgrowth factor receptor MDX-447, or HER2/neu MDX-H210) were shown tocause flu-like symptoms such as fever and chills after the first dose(Curnow, 1997, Cancer Immunol Immunother, 45 210-215). Similar to theeffect of OKT3 on T cells, these antibodies caused a decrease in thenumber of circulating monocytes, which express CD64, and stimulatedincreases in plasma cytokines. A single dose of another monoclonalantibody directed to CD64 (MDX-33) down-modulated the expression of CD64on monocytes and also caused chills, low-grade fever, headache andmuscle aches.

The interaction of T-lymphocytes with antigen-presenting cells (APCs) isone of the initial steps in the activation of an immunological responseto what is perceived by the immune system to be a foreign antigen.Although much attention has been focused on the primary interaction ofthe T-cell receptor with the MHC-antigen on the APC, several other cellsurface components are also involved in T-cell activation. These ligandpairs located on the cell surface of the T-cell and the APC include:LFA-1/ICAM-1 (also ICAM-2 and ICAM-3), CD28/B7, CD2/LFA-3, CD4/MHC ClassII, and CD8/MHC Class I. Interfering with the binding of any of theseligand pairs (e.g., with the use of binding molecules such as monoclonalantibodies) may decrease, inhibit, or discontinue the T-cell responses(de Fourgerolles et al., 1994, J. Exp. Med., 179:619-29; Dustin, M L etal, 1986, J Immunol, 137:245-54).

LFA-1 (consisting of CD11a and CD18 subunits) interaction with ICAM isnecessary for T-cell killing, T-helper and B-cell responses, naturalkilling, and antibody-dependent cytotoxicity. In addition, LFA-1/ICAMinteractions are involved in adherence of leukocytes to endothelialcells, fibroblasts, and epithelial cells, facilitating the migration ofleukocytes from the vasculature to the sites of inflammation (Collins,T., 1995, Science and Medicine, 28-37; Dustin, M L. et al., 1991, AnnualRev Immunology, 9:27-66).

Using antibodies that interfere with LFA-1/ICAM interactions decreasesor inhibits the inflammatory process by blocking the activation ofT-cells and/or the extravasation of leukocytes. In vitro, monoclonalantibodies against LFA-1 or its ligands have inhibited T-cell activation(Kuypers, T. and Roos, D., 1989, Research in Immunology, 140:461-86;Springer, T A, 1987, Annual Rev Immunology, 5:223-52), T-cell dependentB-cell proliferation (Fischer, A. et al., 1986, J Immunol,136:3198-203), target cell lysis (Krensky, A. et al., 1983, J Immunol,131:6711-6), and adhesion of T-cells to vascular endothelium (Dustin, ML. et al., 1988, Journal of Cell Biology, 107:321-31). In mice,anti-CD11a antibodies have induced tolerance to protein antigens(Benjamin, R. et al, 1988, European Journal of Immunology, 18:1079-88;Tanaka, Y. et al., 1995, European Journal of Immunology, 25:1555-8),delayed the onset and reduced the severity of experimental autoimmuneencephalomyelitis (Gordon, E J et al., 1995, Journal of Neuroimmunology,62:153-60), inhibited lupus-associated autoantibody production, andprolonged survival of several types of tissue grafts (Cavazzana-Calco MS, Sarnacki S, Haddad E, et al., Transplantation 1995;59(11):1576-82;Nakakura E K, McCabe S M, Zheng B, Shorthouse R A, et al.,Transplantation 1993;55(2):412-7; Connolly M K, Kitchens E A, Chan B, etal, Clinical Immunology and Immunopathology 1994;72(2):198-203; He Y,Mellon J, Apte R, Niederkorn J., Investigative Ophthalmology and VisualScience 1994;35(8):3218-25; Isobe M, Yagita H, Okumura K, Ihara A.,Science 1992;255:1125-7; Kato Y, Yamataka A, Yagita H, et al., Ann Surg1996;223(1):94-100; Nishihara M, Gotoh M, Fukuzaki T, et al.,Transplantation Proceedings 1995;27(1):372; Talento A, Nguyen M, BlakeT, et al, Transplantation 1993;55(2):418-22; van Dijken P J, Ghayur T,Mauch P, et al., Transplantation 1990;49(5):882-6). In human clinicalstudies, murine anti-CD11a monoclonal antibodies have been shown to helpprevent graft failure following bone marrow transplantation(Cavazzana-Calco M S, Bordigoni P, Michel G, et al., British Journal ofHaematology 1996;93:131-8; Fischer A, Friedrich W, Fasth A., Blood1991;77(2):249-56; Stoppa A M, Maraninchi D, Blaise D, Viens P, et al,Transplant International 1991;4:3-7) and renal transplantation (HourmantM, Le Mauff B, Le Meur Y, et al., Transplantation 1994;58(3):377-80;Hourmant M, Bedrossian J, Durand D, et al., Transplantation1996;62(11):1565-70; Le Mauff B, Hourmant M, Rougier J P, et al.,Transplantation 1991;52(2):291-6). An immunosuppressive drug that couldreduce the incidence of both acute graft rejection and delayed graftfunction, while promoting long-term survival with minimum toxicity withthe potential of tolerance induction would provide major benefits to thefield of renal transplantation.

A need continues to exist for new methods of administering therapeuticcompounds which reduces side effects and which increases theeffectiveness of the therapeutic compound.

SUMMARY OF THE INVENTION

One object of the present invention is to provide an improved method ofadministering a therapeutic compound. This and other objects which willbecome apparent from the following description of enabling embodimentshave been achieved by the method of the invention.

One aspect of the invention is a method for reducing the occurrence offever, headache, nausea and/or vomiting associated with administrationof a therapeutic compound to a mammal in need thereof by administeringto the mammal a first conditioning dose of a non-target cell depletingcompound which binds to a cell surface receptor on a target mammaliancell; and then administering at least a second therapeutic dose of thecompound, wherein the second dose is higher than the first dose.

A further aspect of the invention is a method for treating an LFA-1mediated disorder by administering to a mammal in need thereof a firstconditioning dose of a compound which binds to lymphocyte surfacereceptor LFA-1; and then administering at least a second therapeuticdose of the compound, wherein the second dose is higher than the firstdose. LFA-1 mediated disorders contemplated include psoriasis, asthma,rheumatoid arthritis, multiple sclerosis and transplant rejection. In aspecific embodiment, the graft or transplant is a renal transplant.

A further aspect of the invention is a method for conditioning a mammalto tolerate high doses of a therapeutic compound by administering to themammal a first conditioning dose of a non-target cell depleting compoundwhich binds to a cell surface receptor on a target mammalian cell ; andthen administering at least a second therapeutic dose of the compound,wherein the second dose is higher than the first dose.

Another aspect of the invention is a method for down modulating a cellsurface receptor in a mammalian cell population by contacting a targetmammalian cell displaying a receptor molecule on the surface thereofwith a first dose of a ligand which binds to the receptor molecule anddoes not deplete the mammalian cell population; and then furthercontacting the mammalian cell population with at least a second dose ofthe ligand, wherein the second dose is higher than the first dose.

The following preferred embodiments apply to all the above methods ofthe invention. In preferred embodiments, the therapeutic compoundcomprises a polypeptide which binds to an extracellular domain of thereceptor molecule. A preferred polypeptide is an antibody or a fragmentthereof. In one embodiment, the target mammalian cell is a lymphocytesuch as a T lymphocyte. Non-T cell-depleting compounds or antibodiesthat bind CD11a or CD18 cell surface receptors on a T cell, includingthe humanized anti-CD11a antibody hu1124, are specifically encompassed.

Intravenous or subcutaneous mode of administration of the therapeuticcompound is contemplated. In a specific embodiment, administration isnot more than once per week. In an additional embodiment of each of theabove methods, the methods further comprise administering a thirdtherapeutic dose, wherein the third dose is higher than the second dose.Yet a further embodiment is the administration of a fourth therapeuticdose, wherein the fourth dose is higher than or equal to the third dose.

Also provided is composition comprising an anti-CD11a antibody orfragment thereof, and a pharmaceutical carrier, for use as an activepharmaceutical agent for treating an LFA-1 mediated disorder, whereinthe antibody is a non-target cell-depleting antibody and wherein thecomposition is administered to the mammal as a first conditioning dosefollowed by a second therapeutic dose, wherein the second therapeuticdose is higher than the first dose. Another aspect is the use of acompound which binds to the lymphocyte surface receptor LFA-1 in thepreparation of a medicament for the treatment of an LFA-1 mediateddisorder, which treatment comprises administering to the mammal, a firstconditioning dose of the compound followed by a second therapeutic doseof the compound wherein the second therapeutic dose is higher than thefirst dose.

Other aspects of the invention will become apparent from the followingdescription of preferred embodiments which are not intended to belimiting of the invention.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 shows the dosing schedule used in the study of Example 1. Aindicates time of Physician's Global Assessment and PASI, and skinbiopsy.

FIG. 2 shows the changes in PASI score results by dose group for thestudy of Example 1. It is a summary of Mean Percent Change (+/−SD) inPASI scores from baseline to Days 28 and 56. *Subject 015 withdrewconsent at Day 7 and did not have any data for Days 28 and 56.

FIG. 3 shows the dose response reduction in PASI score results for thestudy of Example 1.

FIGS. 4-A through 4-E show the modulation of lymphocyte CD11a and plasmahu1124 levels for each of Groups A-E, for the study of Example 1.

FIG. 5 summarizes histologic data from Example 1. Histologic evidence ofdecreased ICAM-1, Keratin 16 (Ker 16), and CD11a expression in psoriaticplaques is shown.

FIG. 6 shows the decrease in acute adverse events over time, achieved inExample 1 using the method of the invention. An acute adverse event isone or more of fever (mild), headache (mild), nausea or vomiting within24 hours of dosing. A single patient may have had more than one event.

DEFINITIONS

The term “antibody” is used in the broadest sense and specificallycovers single monoclonal antibodies, antibody compositions withpolyepitopic specificity, as well as antibody fragments (e.g., Fab,F(ab′)₂, scFv and Fv), so long as they exhibit the desired biologicalactivity.

The term “monoclonal antibody” as used herein refers to an antibodyobtained from a population of substantially homogeneous antibodies,i.e., the individual antibodies comprising the population are identicalexcept for possible naturally occurring mutations that may be present inminor amounts. Monoclonal antibodies are highly specific, being directedagainst a single antigenic site. Furthermore, in contrast toconventional (polyclonal) antibody preparations which typically includedifferent antibodies directed against different determinants (epitopes),each monoclonal antibody is directed against a single determinant on theantigen. In addition to their specificity, the monoclonal antibodies areadvantageous in that they are synthesized by the hybridoma culture,uncontaminated by other immunoglobulins. The modifier “monoclonal”indicates the character of the antibody as being obtained from asubstantially homogeneous population of antibodies, and is not to beconstrued as requiring production of the antibody by any particularmethod. For example, the monoclonal antibodies to be used in accordancewith the present invention may be made by the hybridoma method firstdescribed by Kohler & Milstein, Nature, 256:495 (1975), or may be madeby recombinant DNA methods (see, e.g., U.S. Pat. No. 4,816,567 (Cabillyet al.)).

The monoclonal antibodies herein specifically include “chimeric”antibodies (immunoglobulins) in which a portion of the heavy and/orlight chain is identical with or homologous to corresponding sequencesin antibodies derived from a particular species or belonging to aparticular antibody class or subclass, while the remainder of thechain(s) is identical with or homologous to corresponding sequences inantibodies derived from another species or belonging to another antibodyclass or subclass, as well as fragments of such antibodies, so long asthey exhibit the desired biological activity, e.g. binding to a cellsurface receptor (U.S. Pat. No. 4,816,567 (Cabilly et al.); and Morrisonet al., Proc. Natl. Acad. Sci. USA, 81:6851-6855 (1984)).

“Humanized” forms of non-human (e.g., murine) antibodies are chimericimmunoglobulins, immunoglobulin chains or fragments thereof (such as Fv,Fab, Fab′, F(ab′)₂ or other antigen-binding subsequences of antibodies)which contain minimal sequence derived from non-human immunoglobulin.For the most part, humanized antibodies are human immunoglobulins(recipient antibody) in which residues from a complementaritydetermining region (CDR) of the recipient are replaced by residues froma CDR of a non-human species (donor antibody) such as mouse, rat orrabbit having the desired specificity, affinity and capacity. In someinstances, Fv framework residues of the human immunoglobulin arereplaced by corresponding non-human residues. Furthermore, humanizedantibody may comprise residues which are found neither in the recipientantibody nor in the imported CDR or framework sequences. Thesemodifications are made to further refine and optimize antibodyperformance. In general, the humanized antibody will comprisesubstantially all of at least one, and typically two, variable domains,in which all or substantially all of the CDR regions correspond to thoseof a non-human immunoglobulin and all or substantially all of the FRregions are those of a human immunoglobulin consensus sequence. Thehumanized antibody optimally also will comprise at least a portion of animmunoglobulin constant region (Fc), typically that of a humanimmunoglobulin. For further details see: Jones et al., Nature,321:522-525 (1986); Reichmann et al., Nature, 332:323-329 (1988); andPresta, Curr. Op. Struct. Biol., 2:593-596 (1992)).

“Single-chain Fv” or “sFv” antibody fragments comprise the V_(H) andV_(L) domains of antibody, wherein these domains are present in a singlepolypeptide chain. Generally, the Fv polypeptide further comprises apolypeptide linker between the V_(H) and V_(L) domains which enables thesFv to form the desired structure for antigen binding. For a review ofsFv see Pluckthun in The Pharmacology of Monoclonal Antibodies, vol.113, Rosenburg and Moore eds. Springer-Verlag, New York, pp. 269-315(1994).

The term “diabodies” refers to small antibody fragments with twoantigen-binding sites, which fragments comprise a heavy chain variabledomain (V_(H)) connected to a light chain variable domain (V_(L)) in thesame polypeptide chain (V_(H) and V_(L)). By using a linker that is tooshort to allow pairing between the two domains on the same chain, thedomains are forced to pair with the complementary domains of anotherchain and create two antigen-binding sites. Diabodies are described morefully in, for example, EP 404,097; WO 93/11161; and Hollinger et al.,Proc. Natl. Acad Sci. USA 90:6444-6448 (1993).

The expression “linear antibodies” when used throughout this applicationrefers to the antibodies described in Zapata et al. Protein Eng.8(10):1057-1062 (1995). Briefly, these antibodies comprise a pair oftandem Fd segments (V_(H)-C_(H)1-V_(H)-C_(H)1) which form a pair ofantigen binding regions. Linear antibodies can be bispecific ormonospecific.

A “variant” antibody, refers herein to a molecule which differs in aminoacid sequence from a “parent” antibody amino acid sequence by virtue ofaddition, deletion and/or substitution of one or more amino acidresidue(s) in the parent antibody sequence. In the preferred embodiment,the variant comprises one or more amino acid substitution(s) in one ormore hypervariable region(s) of the parent antibody. For example, thevariant may comprise at least one, e.g. from about one to about ten, andpreferably from about two to about five, substitutions in one or morehypervariable regions of the parent antibody. Ordinarily, the variantwill have an amino acid sequence having at least 75% amino acid sequenceidentity with the parent antibody heavy or light chain variable domainsequences, more preferably at least 80%, more preferably at least 85%,more preferably at least 90%, and most preferably at least 95%. Identityor homology with respect to this sequence is defined herein as thepercentage of amino acid residues in the candidate sequence that areidentical with the parent antibody residues, after aligning thesequences and introducing gaps, if necessary, to achieve the maximumpercent sequence identity. None of N-terminal, C-terminal, or internalextensions, deletions, or insertions into the antibody sequence shall beconstrued as affecting sequence identity or homology. The variantretains the ability to bind the receptor and preferably has propertieswhich are superior to those of the parent antibody. For example, thevariant may have a stronger binding affinity, enhanced ability toactivate the receptor, etc. To analyze such properties, one shouldcompare a Fab form of the variant to a Fab form of the parent antibodyor a full length form of the variant to a full length form of the parentantibody, for example, since it has been found that the format of theantibody impacts its activity in the biological activity assaysdisclosed herein. The variant antibody of particular interest herein isone which displays at least about 10 fold, preferably at least about 20fold, and most preferably at least about 50 fold, enhancement inbiological activity when compared to the parent antibody.

The term “down modulating a cell surface receptor” means a process ormethod which reduces the number of molecules of the receptor on thesurface of a cell type relative to the number of molecules of thereceptor before the process or method was performed. For example, theadministration of a therapeutic compound which binds to a cell surfacereceptor will down regulate the receptor if the number of receptormolecules on the surface of the cell is less after administration thanbefore administration of the compound. The number of cell surfacereceptor molecules can be measured histologically using known stainingand counting methods.

A “conditioning dose” is a dose which attenuates or reduces thefrequency or the severity of first dose adverse side effects associatedwith administration of a therapeutic compound. The conditioning dose maybe a therapeutic dose, a sub-therapeutic dose, a symptomatic dose or asub-symptomatic dose. A therapeutic dose is a dose which exhibits atherapeutic effect on the patient and a sub-therapeutic dose is a dosewhich does not exhibit a therapeutic effect on the patient treated. Asymptomatic dose is a dose which induces at least one adverse effect onadministration and a sub-symptomatic dose is a dose which does notinduce an adverse effect.

The term “cell surface receptor” as used herein means any moleculedisplayed on the surface of a cell and available for binding bytherapeutic compounds which contact the surface of the cell. Such a cellsurface molecule is a “receptor” for the therapeutic compound. Humanleukocyte surface markers, including but not limited to CD2, CD29, CD40,CD49a-d and CD58 may be cell surface receptors within the invention.Suitable cell surface receptors also include cell adhesion moleculessuch as the leukocyte integrins CD11a/CD18, CD11b/CD18, CD11c/CD18 andCD11d/CD18 which are heterodimeric surface receptor molecules. Atherapeutic compound may bind to either member of the heterodimericpair. The cell surface receptor may have an extracellular domain whichavailable for binding to a therapeutic compound as well as transmembraneand intracellular domains. Other examples of receptors include proteinkinase receptors capable of intracellular signaling through tyrosinephosphorylation, etc.

The term “non-target cell depleting compound” or a compound or ligandwhich “does not deplete” a cell population means a compound which bindsto a cell surface receptor molecule, but does not substantially reducethe number of cells in the cell population. A non-depleting compound,for example, will reduce the number of cells in a cell population byabout 50% or less, preferably by about 30% or less, more preferably byabout 20% or less, relative to the number of target cells in the cellpopulation prior to contact or treatment with the compound.

The term “LFA-1-mediated disorders” refers to pathological states causedby cell adherence interactions involving the LFA-1 receptor onlymphocytes. Examples of such disorders include T cell inflammatoryresponses such as inflammatory skin diseases including psoriasis;responses associated with inflammatory bowel disease (such as Crohn'sdisease and ulcerative colitis); adult respiratory distress syndrome;dermatitis; meningitis; encephalitis; uveitic; allergic conditions suchas eczema and asthma and other conditions involving infiltration of Tcells and chronic inflammatory responses; skin hypersensitivityreactions (including poison ivy and poison oak); atherosclerosis;leukocyte adhesion deficiency; autoimmune diseases such as rheumatoidarthritis, systemic lupus erythematosus (SLE), diabetes mellitus,multiple sclerosis, Reynaud's syndrome, autoimmune thyroiditis,experimental autoimmune encephalomyelitis, Sjorgen's syndrome, juvenileonset diabetes, and immune responses associated with delayedhypersensitivity mediated by cytokines and T-lymphocytes typically foundin tuberculosis, sarcoidosis, polymyositis, granulomatosis andvasculitis; pernicious anemia; diseases involving leukocyte diapedesis;CNS inflammatory disorder, multiple organ injury syndrome secondary tosepticaemia or trauma; autoimmune haemolytic anemia; myethemia gravis;antigen-antibody complex mediated diseases; all types of transplantationrejection, including graft vs. host or host vs. graft disease; etc.

“Treating” a disease, disorder, condition or cell population includestherapy and prophylactic treatment on an acute short term basis and on achronic long-term basis.

The term “mammal” refers to any animal classified as a mammal, includinghumans, domestic and farm animals, and zoo, sports, or pet animals, suchas dogs, horses, cats, cows, etc. Preferably, the mammal herein ishuman.

A “ligand” or a “compound” which binds to a receptor molecule on thesurface of a cell may be any compound capable of binding to the receptormolecule. The therapeutic compound may, for example, be a small organicmolecule (preferably having a molecular weight of about 1000 g/mole orless) or a polypeptide. Suitable polypeptide compounds or ligands can beprepared by methods known in the art, for example, by isolating peptidesor proteins having high binding affinities for the surface moleculeusing phage display technology. Phage display methods are disclosed, forexample, in U.S. Pat. No. 5,750,373; U.S. Pat. No. 5,821,047; U.S. Pat.No. 5,780,279; U.S. Pat. No. 5,403,484; U.S. Pat. No. 5,223,409; U.S.Pat. No. 5,571,698; etc. In these methods, large libraries of peptidesor proteins displayed on the surface of phage are produced and screenedor panned to select those members of the library which bind strongly toa target molecule which is generally immobilized on a solid support.These methods may be used to select peptides or proteins which bind to acell surface receptor by screening a phage display library using thecell surface receptor as the target molecule. Repeated rounds ofselection and separation of the binders having high binding affinityproduces polypeptide compounds capable of binding to the cell surfacereceptor. The polypeptide may be an antibody or an antibody fragment.

A “therapeutic” compound is any compound which is used in treating amammal.

The term “a compound which binds to lymphocyte surface receptor LFA-1”generally refers to any compound capable of binding to either componentof LFA-1. The compound may be a protein which recognizes and binds toLFA-1, for example a binding protein, an antibody directed againsteither CD11a or CD18 or both, but also includes ICAM-1, soluble forms ofICAM-1 (e.g., the ICAM-1 extracellular domain, alone or fused to animmunoglobulin sequence), antibodies to ICAM-1, and fragments thereof,or other molecules capable of inhibiting the interaction of LFA-1 andICAM-1.

The term “anti-LFA-1 antibody” or “anti-LFA-1 MAb” refers to an antibodydirected against either CD11a or CD18 or both. The anti-CD11a antibodiesinclude, e.g., MHM24 [Hildreth et al., Eur. J. Immunol., 13: 202-208(1983)], R3.1 (IgG1) [R. Rothlein, Boehringer Ingelheim Pharmaceuticals,Inc., Ridgefield, Conn.], 25-3 (or 25.3), an IgG1 available fromImmunotech, France [Olive et al., in Feldmann, ed., Human T cell Clones.A new Approach to Immune Regulation, Clifton, N.J., Humana, 1986 p.173], KBA (IgG2a) [Nishimura et al., Cell. Immunol., 107: 32 (1987);Nishimura et al., ibid, 94: 122 (1985)], M7/15 (IgG2b) [Springer et al.,Immunol. Rev., 68: 171 (1982)], IOT16 [Vermot Desroches et al., Scand.J. Immunol., 33: 277-286 (1991)], SPVL7 [Vermot Desroches et al.,supra], and M17 (IgG2a), available from ATCC, which are rat anti-murineCD11a antibodies. Preferred anti-CD11a antibodies are the humanizedantibodies described in WO 98/23761.

Examples of anti-CD18 antibodies include MHM23 [Hildreth et al., supra],M18/2 (IgG2a) [Sanches-Madrid et al., J. Exp. Med., 158: 586 (1983)],H52 [Fekete et al., J. Clin. Lab Immunol., 31: 145-149 (1990)], Mas191c[Vermot Desroches et al., supra], IOT18 [Vermot Desroches et al.,supra], 60.3 [Taylor et al., Clin. Exp. Immunol., 71: 324-328 (1988)],and 60.1 [Campana et al., Eur. J. Immunol., 16: 537-542 (1986)].

Other examples of suitable LFA-1 binding molecules, includingantibodies, are described in Hutchings et al., supra, WO 91/18011published Nov. 28, 1991, WO 91/16928 published Nov. 14, 1991, WO91/16927 published Nov. 14, 1991, Can. Pat. Appln. 2,008,368 publishedJun. 13, 1991, WO 90/15076 published Dec. 13, 1990, WO 90/10652published Sep. 20, 1990, EP 387,668 published Sep. 19, 1990, EP 379,904published Aug. 1, 1990, EP 346,078 published Dec. 13, 1989, U.S. Pat.No. 5,071,964, U.S. Pat. No. 5,002,869, Australian Pat. Appln. 8815518published Nov. 10, 1988, EP 289,949 published Nov. 9, 1988, and EP303,692 published Feb. 22, 1989.

The antibody is appropriately from any source, including chicken andmammalian such as rodent, goat, primate, and human. Preferably, theantibody is from the same species as the species to be treated, and morepreferably the antibody is human or humanized and the host is human.While the antibody can be a polyclonal or monoclonal antibody,preferably it is a monoclonal antibody, which can be prepared byconventional technology. The antibody is an IgG-1, -2, -3, or -4, IgE,IgA, IgM, IgD, or an intraclass chimera in which Fv or a CDR from oneclass is substituted into another class. The antibody may have an Fcdomain capable of an effector function or may not be capable of bindingcomplement or participating in ADCC.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

It has now been discovered that a dosing schedule in which a firstconditioning dose of a non-target cell depleting compound which binds toa cell surface receptor on a target mammalian cell is followed by asecond higher dose of the compound, is effective in conditioning amammal to tolerate increasing or higher doses of the therapeuticcompound. This dosing schedule allows one to reduce the occurrence ofadverse effects which arise from the initial administration andsubsequent administrations of the therapeutic compound, that is, themethod of the invention reduces the first dose adverse effects ofadministration and condition the mammal to further higher doses.Although some adverse effects such as fever, headache, nausea, vomiting,breathing difficulties, myalgia, chills and changes in blood pressurewill still be observed, the frequency and/or severity of these adverseeffects is surprisingly reduced relative to administration usingconventional dosing schedules such as daily administration of equaldoses of a therapeutic compound. The method of the present invention,therefore, allows one to increase subsequent doses and obtain thetherapeutic benefit of higher doses of the therapeutic compounds, while,at the same time, minimizing the occurrence of adverse side effects.

The therapeutic compound which is administered using the dosing scheduleof the present invention is a non-target cell depleting compound whichbinds to a cell surface receptor on the target cell. Such a compounddoes not substantially reduce the number of cells in the cellpopulation. For example, if the target cell is a lymphocyte,administration of a non-lymphocyte cell depleting compound according tothe dosing schedule of the present invention will result in binding ofthe compound to a lymphocyte cell surface receptor, but will not resultin a decrease in the number of circulating lymphocytes over the courseof administration of the therapeutic compound. It is contemplated thatadministration of the therapeutic compound will, in some cases, resultin increases in the number of target cells in the cell population,however, and such an effect is to be considered within the scope of thepresent invention.

The non-target cell depleting compound which is administered accordingto the method of the invention may be any non-depleting therapeuticcompound which is capable of binding to a cell surface receptor. Manytherapeutic compounds are well known to exert a therapeutic effect bybinding to a selective cell surface marker or receptor. These knowntherapeutic compounds will be apparent to one having ordinary skill inthe art and may be used in the method of the present invention. Suitabletherapeutic compounds include non-peptidic organic compounds, preferablyhaving a molecular weight less than about 1,000 g/mol, more preferablyless than about 600 g/mol; peptide therapeutic compounds, generallycontaining 8 to about 200, preferably about 15 to about 150, morepreferably about 20 to about 100 amino acid residues; and proteintherapeutic compounds, generally having secondary, tertiary and possiblyquaternary structure. Suitable peptides compounds can be prepared byknown solid-phase synthesis or recombinant DNA technology which are wellknown in the art.

A particularly preferred method of selecting a non-depleting peptidecompound is through the use of phage display technology. Using knownphage display methods, libraries of peptides or proteins are prepared inwhich one or more copies of individual peptides or proteins aredisplayed on the surface of a bacteriophage particle. DNA encoding theparticular peptide or protein is within the phage particle. Thesurface-displayed peptides or proteins are available for interaction andbinding to target molecules which are generally immobilized on a solidsupport such as a 96-well plate or chromatography column supportmaterial. Binding and/or interaction of the display peptide or proteinwith a target molecule under selected screening conditions allows one toselect members of the library which bind or react with the targetmolecule under the selected conditions. For example, peptides which bindunder particular pH or ionic conditions may be selected. Alternatively,a target cell population can be immobilized on a solid surface usingknown techniques and the peptide or protein phage library can be pannedagainst the immobilized cells to select peptides or proteins which bindto cell surface receptors on the target cell population. Phage displaytechniques are disclosed, for example, in U.S. Pat. No. 5,750,373; U.S.Pat. No. 5,821,047; U.S. Pat. No. 5,780,279; U.S. Pat. No. 5,403,484;U.S. Pat. No. 5,223,407; U.S. Pat. No. 5,571,698; and others.

One category of preferred polypeptide non-depleting compounds, arecompounds containing an antibody or a fragment thereof whichimmunologically recognize and bind to cell surface receptors. Methods ofpreparing antibodies to specific cell surface receptors are well knownin the art and have been practiced for many years. Suitable antibodiesmay be prepared using conventional hybridoma technology or byrecombinant DNA methods. Preferred antibodies are humanized forms ofnon-human antibodies. Alternatively, antibodies may be prepared fromantibody phage libraries using methods described, for example, in U.S.Pat. No. 5,565,332; U.S. Pat. No. 5,837,242; U.S. Pat. No. 5,858,657;U.S. Pat. No. 5,871,907; U.S. Pat. No. 5,872,215; U.S. Pat. No.5,733,743, and others. Suitable compounds include full-length antibodiesas well as antibody fragments such as Fv, Fab, Fab′ and F (ab′)₂fragments which can be prepared by reformatting the full lengthantibodies using known methods.

Additional preferred polypeptide therapeutic compounds are immunoadhesinmolecules also known as hybrid immunoglobulins. These polypeptides areuseful as cell adhesion molecules and ligands and also useful intherapeutic or diagnostic compositions and methods. An immunoadhesintypically contains an amino acid sequence of a ligand binding partnerprotein fused at its C-terminus to the N-terminus of an immunoglobulinconstant region sequence. A suitable immunoadhesin may contain theextracellular domain of a leukocyte integrin molecule, e.g. LFA-1,LFA-2, LFA-3, MAC-1, p150,95, aDb2, etc., fused to the hinge and CH2and/or CH3 sequences of the a human IgG constant region. Immunoadhesinsand methods of preparing the same are described in U.S. Pat. No.5,428,130; U.S. Pat. No. 5,714,147; U.S. Pat. No. 4,428,130; U.S. Pat.No. 5,225,538; U.S. Pat. No. 5,116,964; U.S. Pat. No. 5,098,833; U.S.Pat. No. 5,336,603; U.S. Pat. No. 5,565,335; etc.

In a method of the invention, adverse side effects are reduced byadministering to a mammal a first conditioning dose of a non-targetcell-depleting compound which binds to a cell surface receptor on atarget mammalian cell followed by a therapeutic dose. The term“therapeutic” in this context means that the compounds binds to thesurface of the target cell and produce a change in the symptoms orconditions associated with the disease or condition which is beingtreated. It is sufficient that a therapeutic dose produce an incrementalchange in the symptoms or conditions associated with the disease; a cureor complete remission of symptoms is not required. One having ordinaryskill in this art can easily determine whether a dose is therapeutic byestablishing criteria for measuring changes in symptoms or conditions ofthe disease being treated and then monitoring changes in these criteriaaccording to known methods. External physical conditions, histologicexamination of affected tissues in patients or the presence or absenceof specific cells or compounds, including in a lesion, associated with adisease may provide objective criteria for evaluating therapeuticeffect. In one example, the method of the invention may be used to treatpsoriasis where therapeutic effect is determined by a physician's globalassessment (PGA) of the patient and by Psoriases Area and Severity Index(PASI) scores. A decrease in PASI score indicates a therapeutic effect.Psoriatic disease activity can also be determined based on OverallLesion Severity (OLS) scale, percentage of total body surface area (BSA)affected by psoriasis, and psoriasis plaque thickness. For asthma, oneindicator of therapeutic effect is a decrease in nonspecific airwayhyperresponsiveness to methacholine challenges (basal and post-allergen;see Examples), upon treatment by the method of the invention. Airwayhyperresponsiveness can be measured by FEV, (volume of air that can beforced from the lungs in 1 second). For transplant or graft survival andfunction, therapeutic effectiveness can be measured, e.g., by theincidence of acute graft rejection, by graft function, and length ofgraft survival, as described in the Examples. Other indicators oftherapeutic effect will be readily apparent to one having ordinary skillin the art and may be used to establish efficacy of the dose.

In the method of the invention, the first dose in followed by a seconddose which is higher than the first dose, that is, contains a greateramount of the therapeutic compound. The first dose serves to conditionthe mammal to tolerate the higher second therapeutic dose. In this way,the mammal is able to tolerate higher doses of the therapeutic compoundthan could be administered initially. Also within the scope of thepresent invention are additional doses, which may be administered afterthe second dose. For example, an additional, third dose which is higherthan or equal to the second dose and an additional fourth dose which ishigher than or equal to the second or third dose are contemplated withinthe method of present invention.

In a further embodiment, the first dose may be repeated one or moretimes before the second higher dose is administered. The first dose maybe administered, for example, one, two or three times, most preferablyonly one time before the second higher dose is administered.

The doses may be administered according to any time schedule which isappropriate for treatment of the disease or condition. For example, thedosages may be administered on a daily, weekly, biweekly or monthlybasis in order to achieve the desired therapeutic effect and reductionin adverse effects. The dosages can be administered before, during orafter the development of the disorder. For example, to prevent hostversus graft or graft versus host rejection, the initial conditioningdose may be administered before, during or after transplantation hasoccurred. The specific time schedule can be readily determined by aphysician having ordinary skill in administering the therapeuticcompound by routine adjustments of the dosing schedule within the methodof the present invention. The time of administration of the first andsecond dosages as well as subsequent dosages will be adjusted tominimize adverse effects while maintaining a maximum therapeutic effect.The occurrence of adverse effects can be monitored by routine patientinterviews and adjusted to minimize the occurrence of side effects, inparticular, fever, headache, nausea and/or vomiting by adjusting thetime of the dosing. Any dosing time is to be considered to be within thescope of the present invention so long as the first conditioning dose ofthe non-depleting compound is administered followed by a second higherdose of the compound. For example, additional doses may be on a daily orweekly schedule followed by subsequent biweekly or monthly doses.

The dosage amount will depend on the specific disease or condition whichis treated and can be readily determined using known dosage adjustmenttechniques by a physician having ordinary skill in treatment of thedisease or condition. The dosage amount will generally lie with anestablished therapeutic window for the therapeutic compound which willprovide a therapeutic effect while minimizing additional morbidity andmortality. Typically, therapeutic compounds will be administered in adosage ranging from 0.001 mg/kg to about 100 mg/kg per dose, preferably0.1-20 mg/kg. The preferred dose of about 0.1-20 mg/kg is particularlyuseful for non-cell depleting compounds containing antibodies orfragments thereof.

Typically, the therapeutic compound used in the method of this inventionis formulated by mixing it at ambient temperature at the appropriate pH,and at the desired degree of purity, with physiologically acceptablecarriers, i.e., carriers that are non-toxic to recipients at the dosagesand concentrations employed. The pH of the formulation depends mainly onthe particular use and the concentration of antagonist, but preferablyranges anywhere from about 3 to about 8. Formulation in an acetatebuffer at pH 5 is a suitable embodiment. Where the therapeutic compoundis an anti-LFA-1 antibody (such as hu1124), a suitable embodiment is aformulation at pH 6.0.

The therapeutic compound, e.g. an anti-LFA-1 antibody, for use herein ispreferably sterile. Sterility can be readily accomplished by sterilefiltration through (0.2 micron) membranes. Preferably, therapeuticpeptides and proteins are stored as aqueous solutions, althoughlyophilized formulations for reconstitution are acceptable.

The therapeutic compound may be formulated, dosed, and administered in afashion consistent with good medical practice. Factors for considerationin this context include the particular disorder being treated, theparticular mammal being treated, the clinical condition of theindividual patient, the cause of the disorder, the site of delivery ofthe agent, the method of administration, the time scheduling ofadministration, and other factors known to medical practitioners. The“therapeutically effective amount” of the therapeutic compound to beadministered will be governed by such considerations, and is the minimumamount necessary to prevent, ameliorate, or treat the disease, forexample an LFA-1-mediated disorder, including treating rheumatoidarthritis, psoriasis, multiple sclerosis, asthma, or prolonging survivalof a transplanted graft. Such amount is preferably below the amount thatis toxic to the host or renders the host significantly more susceptibleto infections.

The therapeutic compound may be administered by any suitable means,including parenteral, subcutaneous, intraperitoneal, intrapulmonary,intranasal, and intralesional administration. Parenteral infusionsinclude intramuscular, intravenous, intraarterial, intraperitoneal, orsubcutaneous administration. Preferably the dosing is given byinjections, most preferably intravenous or subcutaneous injections,depending in part on whether the administration is brief or chronic.

In another aspect of the invention, a method of down-modulating a cellsurface receptor in a cell population in a mammal is provided bycontacting a target mammalian cell displaying a receptor molecule on thesurface thereof with a first dose of a ligand which binds to thereceptor molecule and does not deplete the mammalian cell population;and then further contacting the mammalian cell population with a seconddose of the ligand, wherein the second dose is higher than the firstdose. Binding of the ligand to the cell surface receptor reduces thenumber of receptors on the surface of the cell even though the numbersof cells are not substantially reduced. This reduction in surfacereceptors is associated with reduced side effects such as reduced fever,headache, nausea and/or vomiting.

The non-cell depleting compound or ligand used in this method may besame as described above, and may be provided to the mammal in the samemanner of administration, using the same dosing schedule and in the samedosage amounts as described above.

Another aspect of the invention is the treatment of an LFA-1 mediateddisorder by administering to a mammal, preferably a human patient, inneed of such a treatment a first conditioning dose of a compound whichbinds to lymphocyte surface receptor LFA-1; and administering a secondtherapeutic dose of the compound, where the second dose is higher thanthe first dose. This aspect of the invention is different thanconventional dosing methods for the treatment of such diseases whichgenerally treat with regularly spaced, even doses of a therapeuticcompound. For example, the immunoadhesin LFA-3TIP, which is arecombinant dimeric protein consisting of the first extracellular domainof human LFA-3 fused to the hinge and CH2 and CH3 human IgG regions, hasbeen administered in equal weekly doses of 0.005 mg/kg, 0.025 mg.kg,0.050 mg/kg or 0.075 mg/kg. In a second study, patients received 0.05,0.10 or 0.15 mg/kg in equal doses every four weeks (Krueger et al.). Incontrast, the method of the invention provides a first conditioning doseand then a second therapeutic higher dose.

This method of dose scheduling conditions the patient to tolerate higherdoses of the therapeutic compound than would be tolerated by thepatient, particularly when there are adverse effects on the patients dueto administration of the therapeutic compound. The low first dose of themethod of the invention is useful for reducing the fever, headache,nausea, vomiting, etc. which often accompany an initial administrationof a drug compound. However, it is possible to give a low first dose,within the scope of the invention, to patients who do not experienceadverse effects on first administration.

The first dose conditions the patient to receive a second higher dosewith a reduction in adverse effects which may be observed with higherdoses of therapeutic compounds. Generally, as the dosage amountincreases, the number of adverse effects also increases. The method ofthe invention allows administration of larger therapeutic doses morequickly and with fewer adverse effects. This improves the effectivenessof the therapy since the patient is able to tolerate larger doses andfor a longer time since there are fewer unpleasant side effects.

Any compound which binds to a lymphocyte surface receptor LFA-1 andreduces the severity of symptoms or conditions associated with an LFA-1mediated disease may be used in this embodiment of the invention.Preferred compounds are peptide or protein compounds, more preferablysuch compounds which are or which contain an antibody or fragmentthereof or which are fusions to an antibody fragment such as animmunoadhesin. Particularly preferred compounds are anti-CD11aantibodies or compounds containing fragments thereof.

As described above, the dosage amount will depend on the specific LFA-1mediated disease which is treated and can be readily determined usingknown dosage adjustment techniques by a physician having ordinary skillin treatment of these diseases. The dosage amount will generally liewithin an established therapeutic window for the therapeutic compoundwhich will provide a therapeutic effect while minimizing additionalmorbidity and mortality. Typically, therapeutic compounds will beadministered in a dosage ranging from 0.001 mg/kg to about 100 mg/kg perdose, preferably 0.1-20 mg/kg. The preferred dose of about isparticularly useful for compounds containing antibodies or fragmentsthereof.

The therapeutic compound for treatment of an LFA-1 mediated disease maybe administered by any suitable means, including parenteral,subcutaneous, intraperitoneal, intrapulmonary, intranasal, andintralesional administration. Parenteral infusions includeintramuscular, intravenous, intraarterial, intraperitoneal, orsubcutaneous administration. Preferably the dosing is given byinjections, most preferably intravenous or subcutaneous injections,depending in part on whether the administration is brief or chronic. Asdescribed above in more detail, the therapeutic compound for treatmentof an LFA-1 mediated disease may be formulated, dosed, and administeredin a fashion consistent with good medical practice.

Psoriasis is an inflammatory disease characterized by hyperproliferationof keratinocytes and accumulation of activated T cells in the epidermisand dermis of psoriatic lesions. The upregulation of ICAM-1 onkeratinocytes and its interaction with T-cell LFA-1 in lesional skinindicate that treatment with an anti-CD11a antibody might interfere withthe disease process in psoriasis.

Allergic asthma is characterized by the cardinal features of airwayinflammation, reversible airway obstruction, and hyperresponsiveness. Inpeople with allergic asthma, lymphocytes are believed to play a centralrole in the asthmatic inflammatory response to an inhaled allergen.Chronic asthma symptoms may result from continual activation of lunglymphocytes in response to chronic exposure to perennial allergens(e.g., house dust mite, dog dander, or cat hair) or from sequentialexposure to seasonal allergens to which the patient is reactive.

Within minutes of an inhaled exposure to an aerosolized allergen, thepatient with allergic asthma will experience an immediate response (theearly asthmatic response or EAR), which is characterized by a fall inthe volume of air that can be forced from the lungs in 1 second (FEV₁)over 0-2 hours. In most cases, this fall in FEV₁ is reversible bytreatment with a β-agonist bronchodilator. Approximately 50% of thesepatients will go on to experience a second fall in FEV₁ (the lateasthmatic response or LAR) 3-7 hours after the initial allergenexposure. LAR is associated with more pronounced airway inflammation andincreased bronchial reactivity to nonspecific stimuli. Airwayhyperresponsiveness to the non-allergen-specific challenge ofmethacholine is another cardinal feature of asthma. Hyperresponsivenessof the airways in response to low levels of methacholine is recognizedby a decrease in FEV₁ and may be exacerbated by exposure of the airwaysto allergen, viral infection, or physical irritants. Aerosolizedallergen bronchial challenge, or bronchoprovocation, can be performed inthe laboratory in patients with allergic asthma and is useful andrelevant as a model for the study of anti-asthma medications (Crescioliet al. 1991 Ann Allergy 66:245-51; Cockcroft et al., 1987, Am Rev RespirDis 135:264-267; Ward et al. 1993, Am Rev Respir Dis 147:518-523.). Mostdrugs with proven efficacy in asthma management attenuate airwayresponses (EAR and/or LAR) to inhaled allergens administered in allergenbronchial challenges.

Interleukins (IL-5, IL-3) and granulocyte/macrophage colony stimulatingfactor are known to be important for eosinophil differentiation,maturation, adherence, activation, and degranulation. These cytokinesare produced by T cells isolated from patients with asthma (Walker etal., 1991, J. Immunol. 146:1829-35). The T-cell products IL-4 and IL-13are key mediators of inflammation, increasing IgE levels. Activated Tcells are increased in peripheral blood, bronchoalveolar lavage, andbronchial biopsy specimens from patients with asthma (Azzawi et al.1990, Am Rev Resp Dis 142:1407-13; Corrigan & Kay 1990, Am Rev RespirDis 141:970-7). Lymphocytes are the predominant cell type identified inmorphometric studies of submucosal biopsies obtained from patients withasthma (Djukanovic et al., 1992, Am Rev Respir Dis 145:669-74).

EXAMPLES

The method of the invention is exemplified by the treatment ofpsoriasis, asthma and kidney transplant rejection (LFA-1 mediateddiseases) with a therapeutic compound (an antibody) which binds to acell surface receptor (CD11a). These examples also provide arepresentative example of down modulating a cell surface receptor in amammalian cell population, reducing side effects and of conditioning amammal to tolerate high doses of a therapeutic compound.

Example 1

Phase I Multiple-Dose Dose-Escalation Study of the Effects of hu1124 inSubjects with Moderate to Severe Psoriases. (HUPS249)

The study drug, hu1124, is a known humanized anti-CD11a antibody [see WO98/23761 (humanized MHM24(Fab)-8); Werther W A, et al., Humanization ofan anti lymphocyte function-associated antigen (LFA)-1 monoclonalantibody and re-engineering of the humanized antibody for binding torhesus LFA-1. J Immunol 1996; 157:4986-95). hu1124 is a humanized IgG1version of a murine anti-CD11a monoclonal antibody, MHM24, whichrecognizes human and chimpanzee CD11a. Humanization of MHM24 wasaccomplished by grafting the murine complementarity determining regions(hypervariable region) into consensus human IgG1/κ heavy- andlight-chain sequences. For H and L chain V sequences of hu1124, refer tosequences shown in FIG. 1 in Werther et al., 1996, and in FIGS. 1A and1B in WO 98/23761, incorporated herein by reference.

The study drug, hu1124, was supplied as a single-use, clear, colorless,sterile, non-pyrogenic solution in a glass vial. Each vial contained 10mL of solution at a concentration of 4 mg/mL in 10 mM sodium acetate pH5.0, with 0.02% polysorbate 20, 0.1% sodium acetate trihydrate, and 4%mannitol. No preservative was added to the solution. All study drug wasstored at 2-8° C. (35.6-46.4° F.). The study drug was administered tosubjects by continuous intravenous infusion over 90 minutes into aperipheral vein. The amount of drug to be given was based on subjectweight and the dosage group to which the subject was assigned.

Safety was assessed by adverse events, clinical laboratory assessments,and pre- and post-treatment vital signs. Immunological activity wasmonitored by testing for the effects on cell-mediated immunity reactions(delayed hypersensitivity), tetanus antibody responses, and lymphocytesubpopulations (flow cytometry). Efficacy was monitored by changes inclinical signs and symptoms of the disease (including Psoriasis Area andSeverity Index, PASI scores) and global changes compared with thebaseline condition. Skin biopsies were studied for the effects of hu1124on lymphocytes within psoriatic lesions. Pharmacokinetics of the studydrug were assessed by serial monitoring of plasma samples for hu1124throughout the 98 days following the start of the study.

Thirty-nine subjects with moderate to severe plaque psoriasis wereenrolled at eight study centers. The subjects included Caucasian, Black,Asian and Hispanic subjects. The subjects ranged in age from 26 to 73years, with only one subject older than 70. Subject weight ranged from65 to 122 kg. Between 15 to 72%, overall median of 33%, of the bodysurface area was psoriatic for the subjects enrolled in this study.Baseline PASI scores ranged from 15 to 42, with an overall median of 23for the 39 subjects in this study. Each of the 39 subjects receivedmultiple doses of hu1124, ranging from 0.1-1.0 mg/kg, administered byintravenous infusion. Of the 39 subjects, 10 were enrolled in one of the0.1 mg/kg dose groups (four in the 0.1 mg/kg administered every twoweeks dose group and six in the 0.1 mg/kg/wk dose group), 17 wereenrolled in the 0.3 mg/kg/wk dose group, six were enrolled in the0.3-0.6 mg/kg/wk dose group, and six were enrolled in the 0.3-1.0mg/kg/wk dose group. All but five subjects completed the study. Allsubjects were evaluated for safety and efficacy. The table below showsthe treatment schedule and number of patients in each arm of the study.TABLE 1 Dose Escalation (mg/kg) and Treatment Schedule Day Day Day Day n0 Day 7 14 Day 21 Day 28 35 42 Group A 4 0.1 — 0.1 — 0.1 — 0.1 Group B 60.1 0.1 0.1 0.1 0.1 0.1 0.1 Group C 17 0.3 0.3 0.3 0.3 0.3 0.3 0.3 GroupD 6 0.3 0.4 0.6 0.6 0.6 0.6 0.6 Group E 6 0.3 0.4 0.6 1.0 1.0 1.0 1.0Results—Safety:

The first-dose acute adverse events were primarily fever (reported by17/39 or 44% of subjects), headache (reported by 7/39 or 18% ofsubjects), nausea (reported by 5/39 or 13% of subjects), and vomiting(2/39 or 5% of subjects). Acute (within 48 hours after dosing) adverseevents of fever, headache, or vomiting after the first dose were notreported in subjects who received 0.1 mg/kg every other week or everyweek. The majority of acute adverse events were mild in severity.Importantly, the frequency of acute adverse events decreased withsubsequent doses in each dose group.

Multiple infusions of hu1124 were safe and well-tolerated by thesubjects in this study. The overall decreasing incidence of acuteadverse events after the first dose in each dose group indicates thatthe acute adverse events are the result of initial drug exposure and notabsolute drug level, which in turn indicates an adaptation orconditioning response to the “desensitizing” or “conditioning” dosingschedule of the invention.

Results—Pharmacokinetic/Pharmacodynamics

The mean peak and trough plasma levels of hu1124 appeared to be dosedependent and increased as the dose level of study medication increased.No accumulation of hu1124 was observed in the lower dose groups and asmall degree of accumulation was observed after the maximum doses wereinfused in the higher dose groups. A decrease in CD11a expression wasobserved within 2-4 hours after study medication administration in alldose groups, with full recovery noted before the next dose in the lowerdose groups and within 7-10 days after the hu1124 levels decreased tobelow detection levels. In the two highest dose groups, hu1124 bindingsites remained saturated during the course of treatment.

A reversible increase in the average number of lymphocytes was observedin the higher dose groups after Day 7 with a return to the pretreatmentnumbers after dosing was completed. No effects on the distribution of Tand B subtypes or decreases in T-cell subclasses were observed with anydose.

The tetanus antibody test results indicated that an established humoralantibody response, especially a second set IgG mediated antibodyresponse, is able to persist in the presence of multiple doses ofhu1124. Antibody response was evaluated in 36 of 39 patients by doubleantigen sandwich ELISA. No human anti-hu1124 antibody response wasdetected out to Day 98 after multiple weekly doses.

Results—Efficacy

Efficacy was based on the global assessment of improvement, PASI scores,and histological analysis of skin biopsies. Some clinical improvement(≧poor improvement) was observed in 76% (29/38) of the subjects at Day56. Of the subjects who received at least 0.3 mg/kg/wk, 64% (18/28)experienced clinical improvement of at least fair. Five subjectsexperienced an excellent improvement (i.e., 75-90% improvement frombaseline), with higher rates of clinical improvement noted as the doseof study drug was escalated. Continuous improvement at Day 70 wasobserved in six subjects in the higher dose groups. Subjects infuisedwith the higher doses of hu1124 had greater decreases in their PASIscores compared with subjects infused with the lower doses. Ten subjectshad a 50% decrease in their PASI scores. A dose response as determinedby decreasing PASI scores was noted in the higher dose groups at Day 28;however, by Day 56 no substantial differences were detected among thehigher dose groups. Histological analysis of the skin biopsies revealedsignificant reduction in epidermal thickness and T-cell infiltrationwith clear anti-inflammatory effects and reversal of pathologicalepidermal hyperplasia in subjects treated with at least 0.3 mg/kg/wk.

Conclusions

The conclusions from this study of multiple, escalated infusions ofhu1124 in subjects with moderate to severe plaque psoriasis are asfollows:

-   (a) hu1124 was safe and well-tolerated in subjects who received    multiple, escalated, infused doses ranging from 0.1 mg/kg/every two    weeks to 1.0 mg/kg/wk;-   (b) hu1124 provided clinical improvement in the majority of subjects    as measured by global assessments, PASI scores, and histology;-   (c) hu1124 reduced the CD11a levels on circulating lymphocytes;-   (d) hu1124 did not affect an established humoral antibody response    and did not elicit an immune response;-   (e) hu1124 did not deplete lymphocyte counts; lymphocyte counts    increased during higher dose treatment.

Example 2

A Single-Dose and Multiple-Dose, Escalating Dose Study to Evaluate theSafety, Pharmacokinetics, and Biological Activity of SubcutaneouslyAdministered hu1124 in Subjects with Moderate to Severe Plague Psoriasis(HUPS254)

This study assessed the safety, pharmacokinetics and pharmacodynamics,and biological activity of hu1124 administered by subcutaneous injectionin a single dose and in multiple doses to subjects with moderate tosevere plaque psoriasis.

The study drug, hu1124, was the same as used in Example 1. It wassupplied as single use vials containing 100 milligrams of sterile,pyrogen-free, lyophilized drug product which contains hu1124 antibody ata concentration of 100 mg/mL and 0.02 mmole L-histidine, and 0.96 mmoleβ,β-trehalose, pH 6.0 when reconstituted with 1.0 mL of sterile waterfor injection.

Twenty-six subjects received hu1124 subcutaneously. Subjects in Group A(n=2) received a single injection of 0.3 mg/kg of hu1124. Subjects inGroups B through E (n=24) received eight weekly injections in dosesranging from 0.5-2.0 mg/kg of hu1124.

Table 2 below shows the treatment schedule and number of patients ineach arm of the study. TABLE 2 Dose Escalation and Treatment Schedule nDose (Actual Day Day Day Day Day Day mg/kg Enrollment) Day 0 Day 7 14 2128 35 42 49 Group A (0.3) 2 0.3 — — — — — — — Group B (0.5) 4 0.5 0.50.5 0.5 0.5 0.5 0.5 0.5 Group C 6 0.5 0.7 1.0 1.0 1.0 1.0 1.0 1.0(0.5-1.0) Group D 6 0.7 1.0 1.5 1.5 1.5 1.5 1.5 1.5 (0.7-1.5) Group E 81.0 1.5 2.0 2.0 2.0 2.0 2.0 2.0 (1.0-2.0)

Adverse events were reported and assessed in a manner similar toExample 1. Acute adverse events included headache, fever, chills,myalgia, nausea, and vomiting. Any of these adverse events occurringwithin 48 hours from the time of injection with hu1124 were consideredacute. Safety was assessed by pre- and post-treatment examinations(including vital sign measurements), clinical laboratory assessments(including blood chemistries, hematology, and urinalysis), byexamination of reported adverse clinical events, and by a hearingassessment. Evaluation of efficacy was based on PGA levels ofimprovements, changes in PASI scores, and histological analysis of skinbiopsies at Day 56.

Results—Safety

Subcutaneous administration of hu1124 has been very well tolerated. Nolocal cutaneous reaction has been observed. The incidence of acuteadverse reactions (occurring within 48 hours) that were seen previously,after intravenous administration (Example 1, HUPS249), appeared to havedecreased by approximately 50%. The low incidence of headache (mild) ineight of 26 subjects (31%) and fever (low grade) in two of 26 subjects(8%) was noteworthy. Use of the “desensitizing” or “conditioning” dosingschedule of the invention for dose administration has allowed the safeadministration of up to 2 mg/kg with minimal acute adverse events.

Results—Efficacy

The study population was defined as subjects who had a history of and/orwere considered for systemic therapy for chronic moderate to severeplaque-type psoriasis (BSA>15% and PASI>12) which had been diagnosed forat least six months and had been stable for at least three months.

On the basis of the Physicians' Global Assessment (PGA), and decreasesin Psoriasis Area Severity Index (PASI) score, a clear benefit of hu1124treatment was seen.

Improvement in psoriasis as judged by the PGA suggests a dose response.In Group A, [single dose group (0.3 mg/kg)], no improvement was seen, asexpected. Subjects in Groups B, C, and D showed some improvement at Day56. The improvements were as follows: in Group B (0.5 mg/kg), 1Excellent (75-99% improvement over baseline), 1 Slight (1-24%improvement), 2 Withdrawn; in Group C (0.5-1.0 mg/kg), 2 Good (50-74%improvement), 1 Fair (25-49% improvement), 3 Slight; in Group D (0.7-1.5mg/kg), 1 Good, 1 Fair, 4 Slight. Subjects in Group E show the greatestamount of improvement. In this group, were 1 Excellent, 2 Good, 2 Fair,2 Slight, and 1 Withdrawn. The response in Group E indicates that allsubjects dosed with hu1124 showed some improvement (one subject withdrewat Day 7).

Psoriasis Area and Severity Index (PASI) scores decreased by an averageof 10.9% in the single dose 0.3 mg/kg group, by 47.1% in the 0.5 mg/kggroup, by 36.3% in the 0.5-1.0 mg/kg group, by 33.2% in the 0.7-1.5mg/kg group and by 35.6%, in the 1.0-2.0 mg/kg group. In general, thehigher the PGA, the greater the reduction in PASI score.

This study was later expanded to add 15 more subjects to Group C and 16more to Group E. Thus, Group C had a final enrollment of 21 subjects andGroup E had 24 subjects, adding to a total of 55 subjects enrolled at 10study centers (Group A study was discontinued).

The data generated from the total pool of 55 subjects from the multipledose groups showed the following results and observations. Substantialimprovements in PGA and PASI scores were observed by Day 56. Of the 55subjects in the multiple-dose groups, 45% experienced Good or betterimprovements in PGA (defined as an improvement of ≧50% of all clinicalsigns and symptoms of psoriasis compared to baseline) and 47%experienced at least a 50% decrease in PASI scores. Furthermore, 18% ofthese subjects experienced at least a 75% reduction in PASI scores byDay 56 and were categorized as treatment responders. Among the multipledose groups, higher proportions of subjects in the 0.5-1.0 mg/kg and1.0-2.0 mg/kg groups experienced Good or better improvement in PGA. Ofnote, one subject in the 1.0-2.0 mg/kg group experienced competeresolution of disease symptoms as assessed by PGA. Higher proportions ofsubjects in these two dose groups also experienced at least 50%reductions in PASI scores compared with subjects in the other dosegroups.

Results—Summary

Administration of a subcutaneous formulation of hu1124 waswell-tolerated at the site of injection. Clinical benefits were clearlyseen with Fair, Good, and Excellent responses (PGA) observed in severaldosing groups. The most significant benefits were seen in Group E(1.0-2.0 mg/kg), in particular, where all subjects dosed (one subjectwithdrew) showed improvement as early as Days 28-42 of therapy.

Example 3

An Extended Duration, Multiple-Dose Study to Evaluate the Safety,Pharmacokinetics, and Biological Activity of Intravenously andSubcutaneously Administered hu1124 in Subjects with Moderate to SeverePlague Psoriasis (HUPS256)

In this study, two treatment groups receive multiple intravenous dosesof hu1124 from 0.3 mg/kg to 1.0 mg/kg, for 12 weeks. Each hu1124 dosewill be administered one time weekly over a period of 90 minutes. Threetreatment groups receive multiple subcutaneous doses of hu1124 from 0.7to 4.0 mg/kg, injected one time weekly for 12 weeks.

For intravenous administration, the study drug, hu1124, is supplied as asingle-use, clear, colorless, sterile, non-pyrogenic solution in a glassvial. Each vial contains 10 mL of solution at a concentration of 4 mg/mLin 10 mM sodium acetate pH 5.0, with 0.02% polysorbate 20, 0.1% sodiumacetate trihydrate, and 4% mannitol. No preservative is added to thesolution. All study drug is stored at 2-8° C. (35.6-46.4° F.). Forsubcutaneous administration, the study drug is supplied as single usevials containing 100 milligrams of sterile, pyrogen-free, lyophilizeddrug product which contains hu1124 antibody at a concentration of 100mg/mL and 0.02 mmole L-histidine, and 0.96 mmole β,β-trehalose, pH 6.0when reconstituted with 1.0 mL of sterile water for injection.

Table 3 below shows the treatment schedule and number of patients ineach arm of the study. TABLE 3 Treatment Schedule hu1124 (mg/kg) DAY 0 714 21 28 35 42 49 56 63 70 77 Dosing Schedule (IntravenousAdministration) Group A 0.3 0.3 0.3 0.3 0.3 0.3 0.3 0.3 0.3 0.3 0.3 0.3(n = 6) Group B 0.3 0.6 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 (n = 10)Dosing Schedule (Subcutaneous Administration) Group C 0.7 1.0 1.0 1.01.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 (n = 20) Group D 0.7 2.0 2.0 2.0 2.0 2.02.0 2.0 2.0 2.0 2.0 2.0 (n = 20) Group E 0.7 4.0 4.0 4.0 4.0 4.0 4.0 4.04.0 4.0 4.0 4.0 (n = 20)

Safety is assessed by pre- and post-treatment examinations (includingvital sign measurements), hearing assessments, clinical laboratoryassessments (including blood chemistries, hematology, and urinalysis),antibody to hu1124 (HAHA), and by examination of reported adverseclinical events as in the previous examples. Pharmacokinetics areassessed using various ex vivo immunologic assays. Biological activityis assessed by changes in PASI.

Results

To date, 61 patients have been enrolled and the dosing has been welltolerated at all doses used in the study. One adverse event of localskin reaction has been reported among the 40 patients who have receivedSC doses up to 2.0 mg/kg. None of the 18 patients who have received SCdoses up to 4.0 mg/kg have reported an adverse event of local skinreaction. Clinical benefits are seen with Fair, Good, and Excellentresponses (PGA) observed. To date, anti-CD11a (hu1124) administered bySC injection appears to be safe, well tolerated, and exhibits promisingbiological and clinical activity.

Example 4

Phase III Study of Efficacy, Safety, and Tolerability of SubcutaneousAdministration of hu1124 in Subjects with Moderate to Severe PlaquePsoriasis during Three Phases: First Treatment, Retreatment, andExtended Treatment (ACD2058g)

Subjects receive 12 weekly doses of anti-CD11a (hu1124) or placeboadministered by SC injection, as outlined in the Table of dosingschedule immediately below. The doses consist of an initial conditioningdose at a concentration of 0.7 mg/kg SC and weekly doses of 1.0 mg/kgadministered SC or 2.0 mg/kg SC thereafter. TABLE 4 Dosing Schedule Day0 7 14 21 28 35 42 49 56 63 70 77 Low dose 0.7 1.0 1.0 1.0 1.0 1.0 1.01.0 1.0 1.0 1.0 1.0 (mg/kg) High dose 0.7 2.0 2.0 2.0 2.0 2.0 2.0 2.02.0 2.0 2.0 2.0 (mg/kg)

Anti-CD11a (hu1124) is supplied as in the above Examples 2 and 3. Whenreconstituted with 1.0 mL of Sterile Water for Injection (SWFI), eachvial contains hu1124 at a concentration of 100 mg/mL, as well aspolysorbate 20, L-histidine hydrochloride, and ββ-trehalose, at pH of6.0.

Primary efficacy determinations are made on FT Day 84, which is the endof the First Treatment (FT) period. On FT Day 84, subjects are definedas responders, partial responders, or non-responders according to thefollowing definitions: Responder: PASI has decreased ≧75% from FT Day 0;Partial responder: PASI has decreased ≧50% but <75% from FT Day 0;Non-responder: PASI has decreased <50% from FT Day 0.

This response to therapy determines whether subjects are assigned to theObservation (OB) or Extended Treatment (ET) period after completion ofthe FT Day 84 assessments. Subjects defined as responders enter theObservation (OB) period and are followed either for 6 months or untilrelapse, whichever occurs first. Relapse is defined as loss of 50% ormore of the improvement in PASI achieved between FT Day 0 and FT Day 84(see Section 4.5.3.a). At the time of relapse, subjects who receivedactive drug during the First Treatment (FT) period enter the Retreatment(RT) period and are re-randomized in a 2:1 ratio to anti-CD11a orplacebo, respectively. Subjects who received placebo during the FirstTreatment (FT) period and qualify as responders receive anti-CD11aduring the Retreatment period. Despite re-randomization, subjects remainwithin the dose level group (low dose or high dose) assigned during theFirst Treatment (FT) period. During the Retreatment (RT) period,subjects receive a second course of treatment consisting of 12 weekly SCinjections.

Subjects defined as partial responders or non-responders at the end ofthe First Treatment (FT) period are assigned to the Extended Treatment(ET) period. Subjects remain within the dose levels assigned in theFirst Treatment (FT) period. Subjects who had received anti-CD11a in theFirst Treatment (FT) period are re-randomized 2:1 to anti-CD11a orplacebo, respectively. All subjects who received placebo in the FirstTreatment (FT) period are assigned to anti-CD11a within their doselevel. ET Day 0 occurs on the same day as FT Day 84; hence the twocourses of study drug treatment are continuous over a 24-week period.

In a related study (ACD2062g), subjects who had received prioranti-CD11a treatment and have antibodies to anti-CD11a, and subjects whoare receiving concurrent topical psoriasis therapy or Ultraviolet Blight (UVB) phototherapy are treated on the same dosing regimen as shownin the preceding table.

Efficacy, safety and tolerability are measured as described in the aboveexamples.

Results

The above dosing regimen with initial, lower conditioning dose is welltolerated. Clinical benefits with Fair, Good, and Excellent responses(PGA) are observed.

Example 5

A Phase II Study to Evaluate the Safety and Efficacy of Anti-CD11aantibody, hu1124, in Adults with Allergic Asthma UndergoingAeroallergen-provoked Bronchoconstriction (ACD2017g)

This study evaluates the safety, tolerability, and efficacy of eightweekly SC (subcutaneous) injections of up to 2.0 mg/kg hu1124 in adultsubjects with mild to moderate allergic asthma. The effect of hu1124 onearly and late asthmatic response to an inhaled aeroallergen, and onboth nonspecific and allergen-induced airway hyperresponsiveness, areevaluated. An initial low, conditioning dose of 0.7 mg/kg isadministered followed by subsequent dosing at 2.0 mg/kg.

Thirty-six subjects with allergic asthma, ages 18-60 years, are treatedin this study. Cutaneous responses to allergen (screened by skin pricktesting and assessed by endpoint titration) are measured, andmethacholine bronchial challenges are conducted. These data are used toestimate the starting allergen dose for use in allergen bronchialchallenge, using an empirically derived and validated formula.

During the treatment period, study drug (hu1124 or placebo) isadministered weekly by SC injection, for eight doses over 50 days (onedose per week). The first dose of study drug is administered on Day 0.Subjects undergo allergen bronchial and methacholine challenges.Methacholine challenges, measuring nonspecific airwayhyperresponsiveness (basal and post-allergen), are performed the dayprior to a scheduled allergen challenge and ˜24 hours following thestart of allergen challenge.

The hu1124 antibody is supplied by Genentech as a sterile, pyrogen-free,lyophilized drug product in 10-mL glass vials. When reconstituted with1.0 mL of Sterile Water for Injection (SWFI), each vial contains hu1124at a concentration of 100 mg/mL, 20 mmol of L-histidine, 240 mmol ofβ,β-trehalose, 0.04% polysorbate 20 (pH of 6.0). The placebo formulationhas the same product composition but does not contain hu1124. Study drugis administered by SC injection in the forearm, thigh, or abdomen. Thedosing regimen is shown in the table immediately below. TABLE 5 DosingRegimen Day Treatment 0 7 14 21 28 35 42 49 hu1124 0.7 2.0 2.0 2.0 2.02.0 2.0 2.0 Placebo x x x x x x x xDoses of hu1124 are given in mg/kg.

The safety and efficacy of hu1124 are assessed by the incidence andseverity of adverse events, laboratory tests, physical examinationsincluding vital signs, spirometry, and serum antibody response to hu1124at baseline and during the treatment and follow-up periods. Theincidence and magnitude of changes in hearing are assessed byaudiograms. Efficacy measures include determining the change in LAR andEAR and allergen-induced increase in airway responsiveness. All subjectsare followed for 28 days after dosing is complete.

Blood and urine samples are collected periodically for analysis of bloodchemistries, hematology, and urinalysis. Specific parameters assessedare as follows: Chemistry: sodium, potassium, chloride, bicarbonate,glucose, BUN, creatinine, calcium, phosphorus, magnesium, total anddirect bilirubin, albumin, ALT, AST, alkaline phosphatase, uric acid,total protein. Hematology: CBC with differential and platelet count.Urinalysis: complete urinalysis with microscopic examination. Serumantibodies to hu1124. Serum samples for pharmacokinetic evaluations.Vital sign measurement consists of sitting blood pressure, respiratoryrate, pulse rate, and body temperature measured orally (° C.).

The above dosing regimen is well tolerated and shows efficacy intreating asthma.

Example 6

Phase I/II Study of Anti-CD11a (hu1124) for Renal Transplantation

The use of a non-T cell-depleting, humanized, monoclonal anti-CD11aantibody should cause significantly less toxicity, sensitization, andmore specific immunosuppression than currently available anti-T cellmonoclonal antibodies (OKT3).

This study evaluates the safety, pharmacokinetics and pharmacodynamics(PK/PD), biological activity and clinical effects of 12 weeks ofsubcutaneously administered anti-CD11a (hu1124) at 2 dose levels inpatients undergoing their first cadaveric donor kidney transplant.Patients are treated with 12 doses at weekly intervals and followed for13 weeks. Each dose group has two arms in which standard backgroundimmunosuppression regimens are used, as summarized in Study Design Table5 below. The immunosuppressive agents are cyclosporine, prednisone,mycophenolate mofetil (MMF) and sirolimus. Each patient receives theinitial “conditioning” dose of anti-CD11a (hu1124) at 0.5 mg/kg or 0.7mg/kg, dependent on dose group assignment, on Day 0 at least 1 hourprior to surgery as summarized in Table 6 below. Thereafter, eachpatient receives a maintenance dose of anti-CD11a (hu1124) at 0.5 mg/kgor 2.0 mg/kg each weekly visit for 11 weeks, based on the dose group.TABLE 6 Study Design Dose Group n Group I Anti-CD11a (hu1124) 0.5 mg/kgplus Arm A. Half-dose cyclosporine (2.5-5 mg/kg), sirolimus, prednisone9 Arm B. Full-dose cyclosporine (>5-10 mg/kg), MMF, prednisone 9 GroupII Anti-CD11a (hu1124) 2.0 mg/kg plus Arm A. Half-dose cyclosporine(2.5-5 mg/kg), sirolimus, prednisone 9 Arm B. Full-dose cyclosporine(>5-10 mg/kg), MMF, prednisone 9

TABLE 7 Dosing Regimen Days of Study Dose Group 0 7 14 21 28 35 42 49 5663 70 77 Group I: 0.5 0.5 0.5 0.5 0.5 0.5 0.5 0.5 0.5 0.5 0.5 0.5 0.5mg/kg Group II: 2.0 0.7 2.0 2.0 2.0 2.0 2.0 2.0 2.0 2.0 2.0 2.0 2.0mg/kg

The study drug, anti-CD11a (hu1124), is supplied by XOMA as a sterile,pyrogen-free, lyophilized drug product in 10 mL glass vials which, whenreconstituted with 1.0 mL Sterile Water for Injection (SWFI), containanti-CD11a (hu1124) at a concentration of 100 mg/mL, 20 mmol ofL-histidine, 240 mmol of α,α-trehalose, and 0.04% polysorbate 20 (pH of6.0). Vials are refrigerated at 2° C.-8° C. (36° F.-46° F.).Reconstituted product is stable at room temperature for up to 8 hours.

Safety is assessed by the incidence of adverse events, vital signs,changes in laboratory values compared with baseline (hematology,chemistry, urinalysis), infections, lymphoma, acute rejection episodes,delayed graft function, graft loss, death, incidence of humananti-humanized monoclonal antibody (HAHA).

After initial dosing of the study drug in each patient, thepharmacokinetic characteristics of the study drug are assessed by serialmeasurement of plasma concentrations of anti-CD11a (hu1124) at varioustime points throughout the duration of the study. Flow cytometryanalysis of T cells, T lymphocyte subsets (CD3/CD4/CD8), NK cells, and Blymphocytes, plus expression of CD11a on T lymphocytes, are performed.Gene activation in cytotoxic T lymphocytes (perforin, granzyme B, FAS L)and cytokine gene expression (IL-10, IFN gamma, IL-2, TGF beta, IL-13)in urinary lymphocytes (Vasconcellos et al. 1998, Transplantation 66:562-566) are examined. In addition, the humoral immune response isstudied by examining peripheral blood for anti-HLA antibody production,by ELISA.

The biological activity is measured as follows. Alterations in graftfunction are assessed by serial measurements of serum creatinine, theneed for dialysis during the initial 7 days post transplant, developmentof proteinuria, acute rejection episodes, and response to high-dosesteroid therapy (as measured by serum creatinine). As per transplantcenter standard of care, graft biopsies are obtained at the time ofimplantation, when necessary to confirm the diagnosis of acuterejection, or if rejection does not occur, 1 week after treatmentcompletion (Day 84). Material is obtained for cytokine and cytotoxic Tlymphocyte mRNA analysis, quantitation of interstitial fibrosis, andother potential markers of chronic allograft nephropathy (i.e., collagenexpression).

The above dosing regimen with an initial, lower conditioning dose ofanti-CD11a antibody, followed by a higher therapeutic dose reduces theincidence of both acute graft rejection and delayed graft function, andpromotes long-term survival with minimum toxicity, compared to theadministration of equal doses throughout.

The method of the invention provides a higher therapeutic index thanconventional and current therapy by minimizing toxicity and adverse sideeffects. The foregoing written specification is considered to besufficient to enable one skilled in the art to practice the invention.Indeed, various modifications of the invention in addition to thoseshown and described herein will become apparent to those skilled in theart from the foregoing description and fall within the scope of theappended claims.

1. A method for reducing the occurrence of fever, headache, nauseaand/or vomiting associated with administration of a therapeutic compoundto a mammal in need thereof, comprising: administering to the mammal afirst conditioning dose of a non-target cell-depleting compound whichbinds to a cell surface receptor on a target mammalian cell; andadministering a second therapeutic dose of the compound, wherein thesecond dose is higher than the first dose.
 2. The method of claim 1,wherein the therapeutic compound comprises a polypeptide which binds toan extracellular domain of the receptor molecule.
 3. The method of claim2, wherein the polypeptide is an antibody or a receptor binding fragmentthereof.
 4. The method of claim 1, wherein the target mammalian cell isa lymphocyte.
 5. The method of claim 4, wherein the lymphocyte is aT-cell.
 6. The method of claim 5, wherein the cell surface receptor onthe T cell is CD11a or CD18.
 7. The method of claim 6, wherein the cellsurface receptor is CD11a and the antibody is antibody hu1124.
 8. Themethod of claim 7, wherein the antibody or a receptor binding fragmentthereof is non-lymphocyte depleting.
 9. The method of claim 7, whereinthe antibody is a humanized antibody.
 10. The method of claim 1, furthercomprising administering a third therapeutic dose, wherein the thirddose is higher than the second dose.
 11. The method of claim 10, furthercomprising administering a fourth therapeutic dose, wherein the fourthdose is higher than the third dose.
 12. The method of claim 1, whereinadministration is intravenous or subcutaneous.
 13. The method of claim1, wherein administration is not more than once per week.
 14. The methodof claim 7, wherein the antibody is administered for the treatment ofpsoriasis, asthma, or transplant rejection.
 15. The method of claim 7,wherein the antibody is administered for the treatment of rheumatoidarthritis, systemic lupus erythmatosus or multiple sclerosis.